TAILIEUCHUNG - Báo cáo khoa học: Inhibition of an iron-responsive element/iron regulatory protein-1 complex by ATP binding and hydrolysis

Iron regulatory protein-1 binding to the iron-responsive element of mRNA is sensitive to iron, oxidative stress, NO, and hypoxia. Each of these agents changes the level of intracellular ATP, suggesting a link between iron levels and cellular energy metabolism. | ễFEBS Journal Inhibition of an iron-responsive element iron regulatory protein-1 complex by ATP binding and hydrolysis Zvezdana Popovic and Douglas M. Templeton Laboratory Medicine and Pathobiology University of Toronto Canada Keywords ATP binding ATP hydrolysis energy metabolism iron regulatory proteins ironresponsive element Correspondence D. M. Templeton Laboratory Medicine and Pathobiology University of Toronto 1 King s College Circle Toronto Ontario M5S 1A8 Canada Fax 1 416 978 5959 Tel 1 416 978 3972 E-mail Received 22 February 2007 revised 29 March 2007 accepted 24 April 2007 doi Iron regulatory protein-1 binding to the iron-responsive element of mRNA is sensitive to iron oxidative stress NO and hypoxia. Each of these agents changes the level of intracellular ATP suggesting a link between iron levels and cellular energy metabolism. Furthermore restoration of iron regulatory protein-1 aconitase activity after NO removal has been shown to require mitochondrial ATP. We demonstrate here that the iron-responsive element-binding activity of iron regulatory protein is ATP-dependent in HepG2 cells. Iron cannot decrease iron regulatory protein binding activity in cell extracts if they are simultaneously treated with an uncoupler of oxidative phosphorylation. Physiologic concentrations of ATP inhibit ironresponsive element iron regulatory protein binding in cell extracts and binding of iron-responsive element to recombinant iron regulatory protein-1. ADP has the same effect in contrast to the nonhydrolyzable analog adenosine 5 - P y-imido triphosphate indicating that in order to inhibit iron regulatory protein-1 binding activity ATP must be hydrolyzed. Indeed recombinant iron regulatory protein-1 binds ATP with a Kd of 86 17 pM in a filter-binding assay and can be photo-crosslinked to azido-ATP. Upon binding ATP is hydrolyzed. The kinetic parameters Km pM Vmax nmol-min-1- mg protein -1 are consistent

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